BRCA-1 and BRCA-2 gene mutations have been identified as being responsible for about 90% of all early-onset hereditary breast and ovarian cancers.
BRCA-1 and BRCA-2 gene mutations have also been associated with an increased risk of, or susceptibility to, colon or prostate cancer.
The full sequence of the BRCA-1 and BRCA-2 genes were discovered and/or published in October 1994 and December 1995, respectively. The availability of these sequences and the deduced amino acid sequences has allowed the generation of representative peptide sequences. These peptide sequences have been used as immunogens to raise antibodies capable of detecting specific regions of the BRCA-1 and BRCA-2 proteins.
Approximately 88% of BRCA-1 mutations have been reported to result in the production of a foreshortened protein, from which the terminal end sequences (i.e., carboxy-terminus (C-terminus)) of the BRCA-1 protein are missing. (See Donna Shattuck-Eidens et al., A Collaborative Survey of 80 Mutations . . . , J. Am. Med. Ass'n, Vol. 273 No. 7, Feb. 15, 1995, at 535-541). Data documenting the extent of BRCA-2 mutations, which also result in protein truncation, is available. (See Simon A. Gayther et al., Variation of risks of breast and Ovarian Cancer . . . , BRCA-2 gene, Nature Genetics, Vol. 15, 1997, January 1997, at 103-105). BRCA-1 mutations, however, are reported to have taken the form of single base alterations, single base deletions or multiple base deletions that result in frame shifts that end with downstream stop codons, producing truncation of the BRCA-1 protein consequent to the presence of a gene mutation. As is well known to those skilled in the art, a normal individual carries two unmutated alleles of the BRCA-1 protein while a susceptible individual inherits a normal allele from one parent and a mutated allele from the other parent. These normal and heterozygous allele pairs are carried in every nucleated cell within the body. In tumor tissue, the normal allele has frequently become altered, leaving the susceptible individual with no normal gene copies to produce normal, full-length BRCA-1 protein. In view of the above, it has been hypothesized by the present inventors that the normal nucleated cells of an affected individual will express one normal copy of the BRCA-1 protein and one truncated copy, whereas their tumor cells are likely to possess two altered copies of the protein.
Antibodies capable of detecting the amino-terminus (N-terminus) of the BRCA-1 protein and antibodies capable of detecting the C-terminus of the BRCA-1 protein are known. These antibodies have been used to localize BRCA-1 protein to particular regions of normal cells and tumor cells (see Ralph Scully el al., Location of BRCA-1 in Human Breast Cells and Ovarian Cells, SCIENCE, Vol. 272, Apr. 5, 1996, at 123-124) and, more recently, have been used to monitor expression and localization of protein expressed from transfected constructs containing selected regions of the BRCA-1 protein (see Cindy A. Wilson et al., Differential Subcellular Localization Expression and Biological Toxicity . . . , ONCOGENE, Vol. 14, January 1997, at 1-16).
Antibodies capable of detecting the N-terminus and C-terminus regions of the BRCA-2 protein are also available.
Based upon some of the above-referenced discoveries, Myriad Genetics, Inc. of Salt Lake City, Utah, developed a comprehensive BRCA-1 and BRCA-2 DNA sequence analysis which is marketed under the trade designation BRACAnalysis.TM. comprehensive BRCA-1 and BRCA-2 sequence analysis for susceptibility to breast and ovarian cancer. This genetic analysis reportedly identifies genetic mutations in the entire protein-coding sequences and additional adjacent areas in both the BRCA-1 and BRCA-2 genes. The cost or fee for this genetic analysis is reportedly $2,400.
Due to the complex and comprehensive nature of this DNA-based genetic analysis and the resulting higher cost, it is primarily intended for individuals who are at high risk of hereditary breast or ovarian cancer and those with a diagnosis of breast (especially premenopausal) or ovarian cancer.
Accordingly, there is a need for the development of a less complex and therefore lower cost method for determining cancer susceptibility.
It is therefore the object of the present invention to develop a general screening method for determining susceptibility to breast and ovarian cancer and to other cancers associated with such susceptibility.
It is a more particular object to develop an antibody-based method to detect reduction or loss of the C-terminus regions of BRCA-1 or BRCA-2 protein, which is indicative of a gene mutation.
It is another object of the present invention to develop a test kit for use in conjunction with the developed method.